The present invention relates to methods for controlling Legionella harboring protozoa trophozoites and cysts in aqueous systems. More particularly, the present invention relates to methods for controlling Legionella type bacteria engulphed within a protozoa in the trophozoite form or in Acanthamoeba in the trophozoite and cyst form. The methods of the present invention involve exposing the protozoa to phosphonium salts of the general formula:
(R1)3P+R2.Xxe2x88x92
Wherein R1 is an alkyl group of from 1-8 carbon atoms, R2 is an n-alkyl group having 8-20 carbon atoms and X is an anion consisting of a halide (such as Clxe2x88x92, Brxe2x88x92, etc.), sulfate, nitrate, nitrite etc.
Intracellular bacterial pathogens are a major cause of human morbidity and mortality. Evading hostile intracellular environments is one of the ways pathogens can live within a host cell, even grow within host cells, and yet not be killed or inhibited by the host cell. These parasites have developed ways of interacting and overcoming the host cells natural defense mechanisms. Legionella pneumophila, a bacterium known to cause Legionnaire""s Disease and Pontiac fever in humans is a parasite of this type. While the Legionella cells can be killed while readily exposed to certain chemical agents and antibiotics, Legionella can also be found engulphed (phagocitized) within certain protozoa hosts. Legionella are often found in biofilms adsorbed to solid surfaces in water distribution systems, cooling towers, showers, aquaria, sprinklers, spas, and cleaning baths. Protozoa are natural grazers on surfaces and engulph and digest bacteria as part of their natural life cycle. In most cases, the protozoa digest these bacteria through the use of digestive enzymes in their phagosomes (digestive vacules). In the case of Legionella, however, this is not the case. The protozoa are not readily capable of degrading the engulphed Legionella cells, and in fact, the Legionella grow and increase their numbers while protected within protozoa phagosomes. Legionellosis in humans can be contracted by breathing Legionella aerosols containing either the free-living bacterial cells or by inhaling aerosols of Legionella concentrated within susceptible protozoa. A Legionella control agent, therefore, must be capable of killing free living Legionella, Legionella within protozoa, or the protozoa themselves. The agents described in this invention are capable of killing the free-living Legionella and the host protozoa. Two protozoa species capable of harboring infectious Legionella are Acanthamoeba and Tetrahymena.
In order to effectively control Legionella, in addition to killing the free-living or protozoa an additional factor must be taken into account. Certain protozoa, particularly amoeboid forms have evolved mechanisms for surviving in hostile environments. Examples of hostile environments are high temperature, desiccation, presence of chemical agents/antibiotics, lack of food sources, etc. Upon introduction of a hostile environment, these protozoa revert to a cyst form which is very difficult to kill. The cyst form becomes much less susceptible to chemical agents which readily kill the same organism when in it is in non-cyst (trophozoite) form. Introduction of a chemical control agent to eliminate Acanthamoeba can actually provide the hostile environment to which the protozoa responds by reverting to a cyst form, thereby rendering it invulnerable to the chemical agent. When the cyst contains the pathogen Legionella, the chemical agent can no longer reach the engulphed bacteria, and the chemical treatment is rendered ineffective. As an example, chlorination or bleach is considered essential to control Legionella in water distribution systems. Exposed Legionella are readily killed by low levels of free chlorine (0.2-0.5 xcexcg/ml).
Legionella can also be contained in Acanthamoeba phagosomes if those protozoa are present. The Acanthamoeba sensing the chlorine presence, reverts to a cyst form, inadvertently preserving and protecting the Legionella parasites engulphed within it. Acanthamoeba cysts treated with  greater than 500 times ( greater than 100 xcexcg/ml xe2x80x98freexe2x80x99 chlorine) the concentration needed to kill the trophozoite forms do not kill these cysts. The cysts can revert to the active trophozoite form upon removal of the oxidant. Currently there are no cyst deactivating (killing) agents in commercial use. Control agents that kill the Legionella harboring protozoa cysts would provide a much needed additional tool to safeguard the health of workers and the public against the respiratory pneumonias which can result from inhalation of Legionella or Legionella containing protozoan cysts.